Sur8/Shoc2 Involves Both Inhibition of Differentiation and Maintenance of Self-Renewal of Neural Progenitor Cells via Modulation of Extracellular Signal-Regulated Kinase Signaling

Abstract

Sur8/Shoc2 is a scaffold protein that regulates the Rasextracellular signal-regulated kinase (ERK) pathway. However, the roles of Sur8 in cellular physiologies are poorly understood. In this study, Sur8 was severely repressed in the course of neural progenitor cell (NPC) differentiation in the cerebral cortex of developing rat embryos. Similarly, Sur8 was also critically reduced in cultured NPCs, which were induced differentiation by removal of basic fibroblast growth factor (bFGF). Sur8 regulation occurs at the protein level rather than at the mRNA level as revealed by both in situ hybridization and reverse transcriptase polymerase chain reaction analyses. The role of Sur8 in NPC differentiation was confirmed by lentivirus-mediated Sur8 knockdown, which resulted in increased differentiation, whereas exogenous expression of Sur8 inhibited differentiation. Contrastingly, NPC proliferation was promoted by overexpression, but was suppressed by Sur8 knockdown. The role of Sur8 as an antidifferentiation factor in the developing rat brain was confirmed by an ex vivo embryo culture system combined with the lentivirus-mediated Sur8 knockdown. The numbers and sizes of neurospheres were reduced, but neuronal outgrowth was enhanced by the Sur8 knockdown. The Ras-ERK pathway is involved in Sur8-mediated regulations of differentiation, as the treatment of ERK kinase (MEK) inhibitors blocks the effects of Sur8. The regulations of NPCs' differentiation and proliferation by the Ras-ERK pathway were also shown by the rescues of the effects of bFGF depletion, neuronal differentiation, and antiproliferation by epidermal growth factor. In summary, Sur8 is an antidifferentiation factor that stimulates proliferation for maintenance of self-renewal in NPCs via modulation of the Ras-ERK pathway. STEM CELLS 2011; 29:320-331 Disclosure of potential conflicts of interest is found at the end of this article.

Desmoglein-1/Erbin interaction suppresses ERK activation to support epidermal differentiation

“…It has also been reported to dampen ERK by binding and inhibiting the scaffolding function of SHOC2, which activates ERK by facilitating upstream Ras-Raf interactions (36)(37)(38)(39). However, a potential role for Erbin in regulating ERK signaling in the epidermis has not been addressed.…”

Retinoic-acid-mediated HRas stabilization induces neuronal differentiation of neural stem cells during brain development

“…Most physiological studies of Ras in NSCs focus on the Ras GTPase-activating protein (GAP; e.g. NF1) and its scaffolding protein Sur8 (also known SHOC2) (Moon et al, 2011;Phoenix and Temple, 2010). In addition, most neurological studies of Ras focus on the CNS of adult mice (Kushner et al, 2005;Manabe et al, 2000;Viosca et al, 2009); however, the regulatory mechanism and role of Ras during CNS development is still unclear.…”

“…Primary NSCs were isolated from the telencephalon of C57/BL6 mouse embryos at E14 (Sher et al, 2008) as described previously (Moon et al, 2011). Cells were grown in N2 medium [Dulbecco's Modified Eagle's Medium (DMEM):F12 (1:1) (Gibco, Carlsbad, CA) containing 100 μM putrescine, 30 nM selenite, 20 nM progesterone, 1.55 mg/ml D-(+)-glucose, 25 μg/ml insulin, 0.1 μg/ml apo-transferrin (Sigma-Aldrich), 0.5 mM glutamax, 100 IU/ml penicillin, and 100 μg/ml streptomycin] in the presence of 20 ng/ml bFGF (Peprotech, Princeton, NJ) and 20 ng/ml EGF (Peprotech) in a humidified atmosphere of 95% air and 5% CO 2 at 37°C (Bottenstein and Sato, 1979).…”

“…qRT-PCR analyses were performed in a Rotor-gene Q real-time PCR cycler (Qiagen, Valencia, CA) using SYBR green reagent (Qiagen) with conditions of 95°C for 10 min followed by 40 cycles at 95°C for 5 s and 60°C for 15 s. Relative quantitation of mRNA levels of Tuj1, GFAP, MBP, Nestin, and HRas, KRas and NRas was estimated using the comparative Ct method (ΔΔCt) (Moon et al, 2011). All mRNA values were normalized with respect to GAPDH.…”

“…Ex vivo culture of whole embryo was performed as previously described (Moon et al, 2011). GFP, GFP-HRas, shSc or shHRas lentivirus was mixed with 0.1% Fast Green dye (Sigma-Aldrich), and 1 µl of virus was injected by intraventricular microinjection using a fine glass needle.…”